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Generation of a Primary Hyperoxaluria Type 1 Disease Model Via CRISPR/Cas9 System in Rats

Author(s):

Rui Zheng, Xiaoliang Fang, Lei He, Yanjiao Shao, Nana Guo, Liren Wang, Mingyao Liu, Dali Li* and Hongquan Geng*   Pages 1 - 12 ( 12 )

Abstract:


Background: Primary hyperoxaluria type 1 (PH1) is an inherited disease caused by mutations in alanine-glyoxylate aminotransferase (AGXT). It is characterized by abnormal metabolism of glyoxylic acid in liver leading to endogenous oxalate overproduction and deposition of oxalate in multiple organs, mainly the kidney. Patients of PH1 often suffer from recurrent urinary tract stones, and finally renal failure. There is no effective treatment other than combined liver-kidney transplantation.

Methods: Microinjection were performed to obtain PH1 rats. Urine samples were collected for urine analysis. Kidney tissues were for Western blotting, quantitative PCR, AGT assays and histological evaluation.

Results: In this study, we generated a novel PH1 disease model through CRISPR/Cas9 mediated disruption of mitochondrial localized Agxt gene isoform in rats. Agxt-deficient rats excreted more oxalate in the urine than WT animals. Meanwhile mutant rats exhibited crystalluria and showed a slight dilatation of renal tubules with mild fibrosis in the kidney. When supplied with 0.4% ethylene glycol (EG) in drinking water, mutant rats excreted greater abundance of oxalate and developed severe nephrocalcinosis in contrast to WT animals. Significantly elevated expression of inflammation- and fibrosis-related genes were also detected in mutants.

Conclusion: These data suggest that Agxt-deficiency in mitochondria impairs glyoxylic acid metabolism and leads to PH1 in rats. This rat strain would not only be a useful model for the study of the pathogenesis and pathology of PH1 but also a valuable tool for the development and evaluation of innovative drugs and therapeutics.

Keywords:

Primary hyperoxaluria type 1, nephrocalcinosis, CRISPR/Cas9, kidney, gene editing, mitochondria

Affiliation:

Department of Pediatric Urology, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, Department of Pediatric Urology, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, Department of Pediatric Urology, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, Department of Pediatric Urology, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai



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